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A comparison of single molecule and amplification based sequencing of cancer transcriptomes.


ABSTRACT: The second wave of next generation sequencing technologies, referred to as single-molecule sequencing (SMS), carries the promise of profiling samples directly without employing polymerase chain reaction steps used by amplification-based sequencing (AS) methods. To examine the merits of both technologies, we examine mRNA sequencing results from single-molecule and amplification-based sequencing in a set of human cancer cell lines and tissues. We observe a characteristic coverage bias towards high abundance transcripts in amplification-based sequencing. A larger fraction of AS reads cover highly expressed genes, such as those associated with translational processes and housekeeping genes, resulting in relatively lower coverage of genes at low and mid-level abundance. In contrast, the coverage of high abundance transcripts plateaus off using SMS. Consequently, SMS is able to sequence lower- abundance transcripts more thoroughly, including some that are undetected by AS methods; however, these include many more mapping artifacts. A better understanding of the technical and analytical factors introducing platform specific biases in high throughput transcriptome sequencing applications will be critical in cross platform meta-analytic studies.

SUBMITTER: Sam LT 

PROVIDER: S-EPMC3046973 | biostudies-literature | 2011 Mar

REPOSITORIES: biostudies-literature

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A comparison of single molecule and amplification based sequencing of cancer transcriptomes.

Sam Lee T LT   Lipson Doron D   Raz Tal T   Cao Xuhong X   Thompson John J   Milos Patrice M PM   Robinson Dan D   Chinnaiyan Arul M AM   Kumar-Sinha Chandan C   Maher Christopher A CA  

PloS one 20110301 3


The second wave of next generation sequencing technologies, referred to as single-molecule sequencing (SMS), carries the promise of profiling samples directly without employing polymerase chain reaction steps used by amplification-based sequencing (AS) methods. To examine the merits of both technologies, we examine mRNA sequencing results from single-molecule and amplification-based sequencing in a set of human cancer cell lines and tissues. We observe a characteristic coverage bias towards high  ...[more]

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